Journal
ANALYTICAL CHEMISTRY
Volume 75, Issue 14, Pages 3468-3475Publisher
AMER CHEMICAL SOC
DOI: 10.1021/ac0341057
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- NIDDK NIH HHS [DK 46960] Funding Source: Medline
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Regulated secretion of Zn2+ from isolated pancreatic beta-cells was imaged using laser-scanning confocal microscopy. In the method, beta-cells were incubated in a solution containing the novel fluorescent Zn2+ indicator FluoZin-3. Zn2+ released from the cells reacted with the dye to form a fluorescent product, which was detected by the confocal microscope. The new dye is much brighter than Zinquin, previously used for this application, allowing detection limits of 10-40 nM and temporal resolution of 16 ms/image. The high temporal resolution allowed imaging of isolated fluorescent transients that occurred at the edge of the cells following stimulation with 20 mM glucose or 40 mM K+. Fluorescent transients took 1650 ms to reach a peak from the initial rise and returned to baseline after 170 +/- 50 ms (n = 78 transients from 15 cells). It was concluded that the transients correspond to detection of exocytotic release of Zn2+. Analysis of the temporal and spatial dispersion of the transients indicates that the release of Zn2+ is not diffusion limited but is instead kinetically controlled in agreement with previous observations of insulin release detected by amperometry.
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